This research will develop instrumentation for a fully-automated approach to the evaluation of microregional circulation. This research is based on the working hypothesis that cell permeant fluorescent dyes or "molecular microspheres" can be spatially located by sequential fluorescence imaging of frozen tissue blocks during organ serial sectioning.. The project will develop procedures and adapt instrumentation to evaluate regional flow in an entire organ. During Phase I, feasibility of the technical methodology will be demonstrated through examination of dye optical and extraction properties as well as dye washout and distribution. The validity of the molecular microsphere method will be demonstrated by comparing dye distributions with distributions of 15 microM diameter microspheres simultaneously injected into coronary circulation. Phase I studies will demonstrate feasibility of the working hypothesis and provide design parameters for methods to work with larger hearts and for working with multiple dyes simultaneously. This project will provide researchers with the instrumentation to rapidly and automatically evaluate microregional flow in tissue using procedures similar to the standard procedures involved in radioactive and fluorescent microsphere flow determinations. PROPOSED COMMERCIAL APPLICATIONS Instrumentation and methods to rapidly and automatically evaluate molecular microsphere distributions in tissue will find applications in laboratories engaged in physiology and circulation research. The development of molecular microsphere methodology in the proposal will significantly enhance the commercial market for the Barlow Scientific microsphere imaging system in such laboratories.